Immunoreactive Thomsen-Friedenreich (T) antigen and its precursor, Tn, occur in approximately 90% of all human (adeno) carcinomas (CAs) and in their metastases. In nonCA tissues, these antigens are occluded and inaccessible to the immune system. T & Tn are the immediate precursors of blood type M,N antigens. T & Tn are the first universal CA-diagnostic markers; they are adhesion molecules and persistent autoantigens, from incipience onward; T & Tn's surface densities are often prognostic. Our proposed structures of the T & Tn epitopes based on stepwise degradation and biosynthesis were later confirmed by organic synthesis. The focus will be on Stage I ductal breast CAs. We plan: Study of fresh surgical specimens of primary, preferably Stage I, CAs and tissue-cultured CAs plus control tissues. Full characterization of CA-T & -Tn's structure, location and integration in carrier molecules, macromolecular conformation and distribution in CA. To define extent of T & Tn epitope-clustering, and to determine I & Tn epitopes' interrelationship in CA using metabolically labeled breast CA cells. Immunohistochemical evaluation (peroxidase, gold- silver) with light (ultra)microscopy of spatial distribution of T & Tn epitopes with our anti-T & -Tn rodent monoclonal (Mo) and human monospecific, polyclonal antibodies (Abs). Quantitation of T & Tn epitopes on breast CA cells and on control tissues by RIA and by computerized image analysis, relating T & Tn densities of CA to their ploidy, and correlation of T & Tn densities to invasiveness and autoimmunogenicity of different breast CAs. Lectin and Ab affinity chromatography (C) to separate CA glycoproteins based on their predominant Tn, T and N & M epitopes. Purification of T- & Tn-active glycopeptides obtained by pronase digestion and their reductively beta-eliminated oligosaccharides (OS) by gel filtration, ion-exchange & high-performance liquid C. To determine OS composition by hydrolysis, enzymatically, colorimetrically and by gas C (GC) of alditol acetates, and structures using glycosidases, permethylation, GC-mass spectrometry (GC/MS) & NMR. Amino acid sequencing will be by subtractive Edman degradation. To isolate guinea pig L-lO hepatoCA Tn-, T- and N- & M-active glycolipids (GLs) on s large scale, and define the carbohydrate structures of the T- & Tn-specific GLs. Study of human CAs and lymphomas for expression and nature of T- & Tn-active GLs; relate findings to degree of malignancy. Immunostaining with our MoAbs to locate and identify active GLs. To analyze OS of active L-lO GLs by GC/MS after trifluoroacetolysis, reduction and permethylation.